It is proposed to purify the cytochrome P-450 used to cleave the side-chain of cholesterol from mitochondria of mouse adrenal tumor cells (Y-1) and to characterize the enzyme with respect to physical and enzymatic properties for comparison with the enzyme from beef and that from normal mouse adrenal. The binding characteristics of cholesterol, 20S-hydroxycholesterol and 20s, 22R-dihydroxycholesterol to the enzyme will be examined. The possible role of microfilaments and microtubules in the responses of tumor cells to ACTH will be examined. In the case of microfilaments, fluorescent-labeled heavy meromyosin will be used to decorate filaments within tumor cells incubated with and without ACTH and cyclic AMP. Attempts will be made to purify the tumor cell actin and the colchicin-binding protein (? tubulin) observed in adrenal cells. The durations of the various phases of the cell cycle will be studied; the possible influence of ACTH on the cycle and vice-versa -the possible influence of the cycle on the response of tumor cells to ACTH, will also be examined. The amounts and if possible the synthesis of two steriodogenic enzymes (side-chain cleavage P-450 and 20 alpha-hydroxy-steroid dehydrogenase) will be examined at various phases of the cycle. The processes by which the steroidogenic capacity of the cells is enhanced by injection of tumor cells into mice will be investigated. Two aspects of this process will be examined, namely changes in the cholesterol side-chain cleavage enzyme system as the cells grow into tumors within the host and the possible contributions of host endocrine glands and plasma to this process.